The present invention provides novel hydroxypyrrolidine compounds, their use as medicaments, compositions containing them and processes for their preparation.
Platelet adhesion and aggregation are initiating events in arterial thrombosis. Although the process of platelet adhesion to the sub-endothelial surface may have an important role to play in the repair of damaged vessel walls, the platelet aggregation that this initiates can precipitate acute thrombotic occlusion of vital vascular beds, leading to events with high morbidity such as myocardial infarction and unstable angina. The success of interventions used to prevent or alleviate these conditions, such as thrombolysis and platelet-mediated occlusion or re-occlusion also compromises angioplasty.
A number of converging pathways lead to platelet aggregation. Whatever the initial stimulus, the final common event is a cross-linking of platelets by binding of fibrinogen to a membrane-binding site, glycoprotein IIb/IIIa (GPIIb/IIIa). The high anti-platelet efficacy of antibodies or antagonists for GPIIb/IIIa is explained by their interference with this final common event. However, this efficacy may also explain the bleeding problems that have been observed with this class of agent. Thrombin can produce platelet aggregation largely independently of other pathways but substantial quantities of thrombin are unlikely to be present without prior activation of platelets by other mechanisms. Thrombin inhibitors such as hirudin are highly effective anti-thrombotic agents, but again may produce excessive bleeding because they function as both anti-platelet and anti-coagulant agents (The TIMI 9a Investigators (1994), Circulation 90, pp. 1624-1630; The Global Use of Strategies to Open Occluded Coronary Arteries (GUSTO) IIa Investigators (1994) Circulation 90, pp. 1631-1637; Neuhaus K. L. et. al. (1994) Circulation 90, pp. 1638-1642).
It has been found that ADP acts as a key mediator of thrombosis. ADP-induced platelet aggregation is mediated by the P2T receptor subtype located on the platelet membrane. The P2T receptor (also known as P2YADP or P2TAC) is primarily involved in mediating platelet aggregation/activation and is a G-protein coupled receptor. The pharmacological characteristics of this receptor have been described, for example, in the references by Humphries et al., Br. J. Pharmacology, (1994). 113, 1057-1063, and Fagura et al., Br. J. Pharmacology (1998) 124, 157-164. Recently it has been shown that antagonists at this receptor offer significant improvements over other anti-thrombotic agents (see J. Med. Chem. (1999) 42, 213). There is a need to find P2T (P2YADP or P2TAC) antagonists as anti-thrombotic agents.
In a first aspect the invention provides a compound of formula (I): 
wherein:
R1 is H, CH2R5 or COR6;
R2 is alkyl C1-6 or alkenyl C1-6, optionally substituted by one or more groups selected from alkyl C1-6, halogen;
R3 is cycloalkyl C3-8, optionally substituted by R7;
R4 is H or alkyl C1-6, optionally substituted by one or more halogens;
R5 is H, phenyl or alkyl C1-6, optionally substituted by halogen, OR8, phenyl;
R6 is OR9 or alkyl C1-6, optionally substituted by one or more groups selected from halogen, OR10, phenyl;
R7 is phenyl, optionally substituted by one or more groups selected from alkyl C1-6, halogen, OR8;
R8, R9 and R10, are independently H or alkyl C1-6, optionally substituted by one or more groups selected from halogen or alkyl C1-6;
or a pharmaceutically acceptable salt or solvate thereof, or a solvate of such a salt.
Preferably the compound of formula (I) has the following stereochemistry: 
Where R3 is 
the stereochemistry is preferably 
Preferably R1 is H, CH2Ph, CH2CH2OH, or CO2tBu.
Preferably R2 is n-Pr.
Preferably R3 is cycloalkyl C3-8 substituted by phenyl.
Preferably R4 is H or methyl.
Compounds of the invention include:
[3R-[3xcex1,4xcex2(1R*,2S*)]]-4-[7-[(2-Phenylcyclopropyl)amino]-5-(propylthio)-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-yl]-3-pyrrolidinol;
[3S-[3xcex1,4xcex2(1S*,2R*)]]-3-Hydroxy-4-[7-[(2-phenylcyclopropyl)amino]-5-(propylthio)-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-yl]-1-pyrrolidinecarboxylate, 1,1-dimethylethyl ester,
[3S-[3xcex1,4xcex2(1R*,2S*)]]-3-Hydroxy-4-[7-[(2-phenylcyclopropyl)amino]-5-(propylthio)-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-yl]-1-pyrrolidinecarboxylate, 1,1-dimethylethyl ester,
[3S-[3xcex1,4xcex2(1S*,2R*)]]-4-[7-[(2-Phenylcyclopropyl)amino]-5-(propylthio)-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-yl]-3-pyrrolidinol;
[3R-[3xcex1,4xcex2(1R*,2S*)]]-4-[7-[N-Methyl-N-(2-phenylcyclopropyl)amino]-5-(propylthio)-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-yl]-3-pyrrolidinol,
[3R-[3xcex1,4xcex2(1R*,2S*)]]-1-Hydroxyethyl-4-[7-[(2-phenylcyclopropyl)amino]-5-(propylthio)-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-yl]-3-pyrrolidinol;
[3R-[3xcex1,4xcex2(1R*,2S)]]-4-[7-[(2-Phenylcyclopropyl)amino]-5-(propylthio)-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-yl]-1-(phenylmethyl)-3-pyrrolidinol;
[3R-[3xcex1,4xcex2(1R*,2S*)]]-1-Acetyl-4-[7-[(2-phenylcyclopropyl)amino]-5-(propylthio)-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-yl]-3-pyrrolidinol.
or a pharmaceutically acceptable salt or solvate thereof, or a solvate of such a salt.
The invention further provides a process for the preparation of a compound of formula (I) which comprises:
a. For compounds of formula (I) where R1 is H, reacting a compound of formula (II): 
wherein R2 is as defined above and P is a protecting group, preferably t-BuOCO, with R3R4NH, wherein R3 and R4 are as defined in (I), and a base, preferably triethylamine or N,N-diisopropylethylamine, in the presence of an inert solvent preferably acetonitrile, preferably at a temperature between about 20xc2x0 C. and about 100xc2x0 C. and optionally thereafter removing any protecting groups.
Examples of protecting groups include t-BuOCO and CH2Ph. Protecting groups can be added and removed using known reaction conditions. The use of protecting groups is fully described in xe2x80x98Protective Groups in Organic Chemistryxe2x80x99, edited by J W F McOmie, Plenum Press (1973), and xe2x80x98Protective Groups in Organic Synthesisxe2x80x99, 2nd edition, T W Greene and P G M Wutz, Wiley-Interscience (1991).
A compound of formula (II) can be prepared by diazotizing a compound of formula (III): 
where R2 and P are defined above, and where necessary other reactive groups might also be protected, with a C1-6 alkyl nitrite, preferably iso-amylnitrite in the presence of an inert solvent preferably acetonitrile at a temperature of between about 20 and about 80xc2x0 C., or with an alkali metal nitrite, preferably sodium nitrite, under aqueous acidic conditions, preferably aqueous hydrochloric or acetic acid and preferably at a temperature between about 0xc2x0 C. and about 20xc2x0 C.
A compound of formula (III) can be prepared by reacting a compound of formula (IV): 
wherein P is a protecting group, with a compound of formula (V): 
wherein R2 is as defined in formula (I) and is preferably n-propyl. The reaction is carried out in the presence of a base, preferably triethylamine or N,N-diisopropylethylamine, in an inert solvent preferably N,N-dimethylformamide or n-butanol, at a temperature between about 100xc2x0 C. and about 150xc2x0 C.
The preparation of the formula (IV) racemate is described in Okada et al., Chem. Pharm. Bull. (1993), 41, 132-8; the preparation of formula (IV) enantiomers is described in Schaus, et al., J. Org. Chem. (1997), 62, 4197-9; the preparation of a compound of formula V (R2 is n-propyl) is described in EP 508687.
Compounds of formula (I) where R2 is other than n-propyl are prepared by displacement of the sulphone group from a compound of formula (VI): 
where R2 is n-propyl, P, R3 and R4 are defined above, using either a sodium alkylthiolate (R2SNa) in the presence of an inert solvent, preferably N,N-dimethylformamide, preferably at a temperature between about 0xc2x0 C. and about 50xc2x0 C. or sodium hydrosulphide (NaSH, in the presence of an inert solvent preferably N,N-dimethylformamide. The latter reaction is followed by alkylation with an alkyl halide (R2X, where X is a leaving group preferably bromide or iodide), preferably at a temperature between about 0xc2x0 C. and about 50xc2x0 C. and optionally thereafter removing any protecting groups.
The preparation of the compound of formula (VI), where R2 is n-propyl, is preferably carried out by reacting a compound of formula (I), where R1 has been protected as described above, with a peracid, preferably m-chloroperbenzoic acid, in the presence of an inert chlorocarbon solvent such as dichloromethane or a mixture of dichloromethane and methanol, at a temperature between about 0xc2x0 C. and about 50xc2x0 C.
b. For compounds of formula (I) where R1 is CH2R5, where R5 is defined in formula (I), the reaction scheme outlined in a above is followed by reductive amination using an aldehyde (R5CHO) and a reducing agent, preferably sodium triacetoxyborohydride, and optionally thereafter removing any protecting groups. The reductive amination reaction is preferably carried out in the presence of an inert solvent preferably N,N-dimethylformamide, tetrahydrofuran or a mixture of acetonitrile and N-methylpyrrolidone and preferably at a temperature between about 0xc2x0 C. and about 50xc2x0 C.
c. For compounds of formula (I) where R1 is COR6, where R6 is defined in formula (I), the reaction scheme outlined in a above is followed by acylation using an acid halide (R6COX) or anhydride ((R6CO)2O) or an acid (R6CO2H) in the presence of a suitable activating agent preferably N,Nxe2x80x2-carbonyldiimidazole or N,Nxe2x80x2-dicyclohexylcarbodiimide, and a base preferably triethylamine or N,N-diisopropylethylamine, and optionally thereafter removing any protecting groups. The acylation is preferably carried out in the presence of an inert solvent preferably dichloromethane, chloroform or tetrahydrofuran and preferably at a temperature between about 0xc2x0 C. and about 50xc2x0 C.
Compounds of formula (II), (III), (IV) and (V) form a further aspect of the invention.
Salts of the compounds of formula (I) may be formed by reacting the free base, or a salt or a derivative thereof, with one or more equivalents of the appropriate acid (for example a hydrohalic (especially HCl), sulphuric, oxalic or phosphoric acid). The reaction may be carried out in a solvent or medium in which the salt is insoluble or in a solvent in which the salt is soluble, e.g. water, ethanol, tetrahydrofuran, or diethyl ether, which may be removed in vacuo or by freeze drying. The reaction may also be a metathetical process or it may be carried out on an ion exchange resin. The non-toxic physiologically acceptable salts are preferred, although other salts may be useful, e.g. in isolating or purifying the product.
The compounds of the invention act as P2T (P2YADP or P2TAC) receptor antagonists. Accordingly, the compounds are useful in therapy, including combination therapy, particularly they are indicated for use as: inhibitors of platelet activation, aggregation and degranulation, promoters of platelet disaggregation, anti-thrombotic agents or in the treatment or prophylaxis of unstable angina, coronary revascularisation procedures including angioplasty (PTCA), myocardial infarction, perithrombolysis, primary arterial thrombotic complications of atherosclerosis such as thrombotic or embolic stroke, transient ischaemic attacks, peripheral vascular disease, myocardial infarction with or without thrombolysis, arterial complications due to interventions in atherosclerotic disease such as angioplasty, endarterectomy, stent placement, coronary and other vascular graft surgery, thrombotic complications of surgical or mechanical damage such as tissue salvage following accidental or surgical trauma, reconstructive surgery including skin and muscle flaps, conditions with a diffuse thrombotic/platelet consumption component such as disseminated intravascular coagulation, thrombotic thrombocytopaenic purpura, haemolytic uraemic syndrome, thrombotic complications of septicaemia, adult respiratory distress syndrome, anti-phospholipid syndrome, heparin-induced thrombocytopaenia and pre-eclampsia/eclampsia, or venous thrombosis such as deep vein thrombosis, venoocclusive disease, haematological conditions such as myeloproliferative disease, including thrombocythaemia, sickle cell disease; or in the prevention of mechanically-induced platelet activation in vivo, such as cardio-pulmonary bypass and extracorporeal membrane oxygenation (prevention of microthromboembolism), mechanically-induced platelet activation in vitro, such as use in the preservation of blood products, e.g. platelet concentrates, or shunt occlusion such as in renal dialysis and plasmapheresis, thrombosis secondary to vascular damage/inflammation such as vasculitis, arteritis, glomerulonephritis, inflammatory bowel disease and organ graft rejection, conditions such as migraine, Raynaud""s phenomenon, conditions in which platelets can contribute to the underlying inflammatory disease process in the vascular wall such as atheromatous plaque formation/progression, stenosis/restenosis and in other inflammatory conditions such as asthma, in which platelets and platelet-derived factors are implicated in the immunological disease process. Further indications include treatment of CNS disorders and prevention of the growth and spread of tumours.
In particular, the compounds of the invention are useful in the treatment or prevention of myocardial infarction, thrombotic stroke, transient ischaemic attacks, peripheral vascular disease and stable and unstable angina, especially unstable angina.
The invention also provides a method of treatment or prevention of the above disorders which comprises administering to a patient suffering from or susceptible to such a disorder a therapeutically effective amount of a compound according to the invention.
According to the invention there is further provided the use of a compound according to the invention as an active ingredient in the manufacture of a medicament for use in the treatment or prevention of the above disorders.
The compounds may be administered topically, e.g. to the lung and/or the airways, in the form of solutions, suspensions, HFA aerosols and dry powder formulations; or systemically, e.g. by oral administration in the form of tablets, pills, capsules, syrups, powders or granules, or by parenteral administration in the form of sterile parenteral solutions or suspensions, by subcutaneous administration, or by rectal administration in the form of suppositories or transdermally.
The compounds of the invention may be administered on their own or as a pharmaceutical composition comprising the compound of the invention in combination with a pharmaceutically acceptable diluent, adjuvant or carrier. Particularly preferred are compositions not containing material capable of causing an adverse, e.g. an allergic, reaction.
Dry powder formulations and pressurised HFA aerosols of the compounds of the invention may be administered by oral or nasal inhalation. For inhalation the compound is desirably finely divided. The compounds of the invention may also be administered by means of a dry powder inhaler. The inhaler may be a single or a multi dose inhaler, and may be a breath actuated dry powder inhaler. One possibility is to mix the finely divided compound with a carrier substance, e.g. a mono-, di- or polysaccharide, a sugar alcohol or another polyol. Suitable carriers include sugars and starch. Alternatively the finely divided compound may be coated by another substance. The powder mixture may also be dispensed into hard gelatine capsules, each containing the desired dose of the active compound. Another possibility is to process the finely divided powder into spheres which break up during the inhalation procedure. This spheronized powder may be filled into the drug reservoir of a multidose inhaler, e.g. that known as the Turbuhaler(copyright) in which a dosing unit meters the desired dose which is then inhaled by the patient. With this system the active compound with or without a carrier substance is delivered to the patient.
The pharmaceutical composition comprising the compound of the invention may conveniently be tablets, pills, capsules, syrups, powders or granules for oral administration; sterile parenteral or subcutaneous solutions, suspensions for parenteral administration or suppositories for rectal administration.
For oral administration the active compound may be admixed with an adjuvant or a carrier, e.g. lactose, saccharose, sorbitol, mannitol, starches such as potato starch, corn starch or amylopectin, cellulose derivatives, a binder such as gelatine or polyvinylpyrrolidone, and a lubricant such as magnesium stearate, calcium stearate, polyethylene glycol, waxes, paraffin, and the like, and then compressed into tablets. If coated tablets are required, the cores, prepared as described above, may be coated with a concentrated sugar solution, which may contain e.g. gum arabic, gelatine, talcum, titanium dioxide, and the like.
Alternatively, the tablet may be coated with a suitable polymer dissolved either in a readily volatile organic solvent or an aqueous solvent.
For the preparation of soft gelatine capsules, the compound may be admixed with e.g. a vegetable oil or polyethylene glycol. Hard gelatine capsules may contain granules of the compound using either the above mentioned excipients for tablets, e.g. lactose, sacharose, sorbitol, mannitol, starches, cellulose derivatives or gelatine. Also liquid or semisolid formulations of the drug may be filled into hard gelatine capsules.
Liquid preparations for oral application may be in the form of syrups or suspensions, for example solutions containing the compound, the balance being sugar and a mixture of ethanol, water, glycerol and propylene glycol. Optionally such liquid preparations may contain colouring agents, flavouring agents, saccharine and carboxymethylcellulose as a thickening agent or other excipients known to those skilled in art.